Vitamin D-3 induces the de novo synthesis of calmodulin in Phaseolus vulgaris root segments growing in vitro

Abstract

Vitamin D-3 affects growth and Ca 2+ transport in Phaseolus vulgaris roots by a mechanism dependent on de novo protein synthesis. The objective of this work was to identify the protein(s) induced by the sterol. Phaseolus vulgaris root segments cultured in vitro were used. Protein extracts of control cultures and cultures treated with 10 −9 M vitamin D-3 for 2 h labelled with [ 14C]leucine and [ 3H]leucine, respectively, were mixed and electrophoresed on sodium dodecyl sulfate polyacrylamide gels. Examination of 3H: 14C ratios in slices of gels revealed that vitamin D-3 stimulates production of a protein which exhibited Ca 2+-dependent electrophoretic mobility. The apparent molecular weight was 14500 in the presence of 1 mM Ca 2+ and 18 000 in the presence of 5 mM EGTA. This protein was heat- and acid-stable, bound 45Ca 2+ and had an isoelectric pH of 3.8-4.1. These data are consistent with the properties of calmodulin. In agreement with these observations, higher levels of calmodulin were detected in roots treated with vitamin D-3 than in control roots by means of assays based on the activation of calmodulin-dependent enzymes. Moreover, estimation of calmodulin relative synthesis and degradation rates in roots cultures in the absence and presence of vitamin D-3 indicated that the sterol increases calmodulin synthesis without affecting its degradation. In addition, the results suggest that the synthesis of other low-molecular-weight Ca 2+-binding proteins may be affected by vitamin D-3.